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. Author manuscript; available in PMC: 2023 Apr 27.
Published in final edited form as: Nat Biomed Eng. 2022 Apr 27;6(4):463–475. doi: 10.1038/s41551-022-00880-8

Fig. 8. Kinesin-2 knockout hPSCs and organoids reveal secretion defects in disease-relevant signaling pathways.

Fig. 8.

(a) Schematic of EV purification and analysis experiment. (b) Representative immunoblots of endogenous ciliary proteins (green type) in supernatants (SN) and lysates (LY) from control and KIF3A−/− hPSCs. β-actin is shown as a loading control. (c) Immunoblots of supernatants showing key components of the PKD (blue type) and hedgehog (red type) pathways in these supernatants. β-actin, β-catenin, and flotillin (FLOT1) are shown as loading controls. (d) A panel of non-ciliary EV components in supernatants. (e) PC1 expression in the supernatants of kidney organoids in suspension. Three control and two KIF3A−/− lines are shown. (f) Quantification of PC1 in EVs from control and mutant hPSCs (mean ± s.e.m., n ≥ 6 biological replicates per condition, total of 14 distinct cell lines. ***, p < 0.001). (g) Quantification of IFT88, PC1, and PTCH1 in EVs from control and mutant hPSCs (n = 4 biological replicates, p < 0.01). (h) Representative immunoblots of EV from control, PKD1−/−, and kinesin-2 knockout hPSCs, ± SAG treatment, with (i) quantification (mean ± s.e.m., n = 3 biological replicates per condition, total of 6 distinct cell lines).